CH2210

Amino Acid Metabolism

Fate of Dietary ProteinDietary protein

Denatured and partially hydrolyzed protein (large polypeptides)

Stomach: HCl, pepsin

Amino acids and dipeptides

small intestine: proteases

Amino acids in bloodstream

intestinal lining: proteases

Overview of Amino Acid Catabolism

H3N C C

H

RO

OThe catabolism of amino acids takes

place in three stages:

1) Removal of the amino group, leaving the carbon

skeleton of the amino acid.

2) Breakdown of the carbon skeletons to a glycolytic

intermediate, citric acid cycle intermediate, or acetyl-S-CoA.

3) Oxidation of these intermediates to CO2 and H2O

with the production of ATP.

small pieces

CO2 H2O

ATP

aspartate oxaloacetateNH3

C C

H

O

O

CH2C

O

OO

C CO

O

CH2C

O

O

NH4+

glutamine

α-ketoglutarateNH3

C C

H

O

OCH2C

O

H2NCH2

O

C CO

OCH2C

O

OCH2

2 NH4+

alanine pyruvateNH3

C C

H

H3CO

O

O

C CH3CO

ONH4+

Representative Pathways of Amino Acid Catabolism

phenylalanine

fumarate

acetoacetate

NH3

C C

H

CH2O

OC

CCCC C

CH2 CCO

OH3C

O

CH CCHO

OC

O

O

HH

H

H H

NH4+CO2

+

citrateoxaloacetate

fumarate

succinate

succinyl-CoA

malate isocitrate

aconitate

α-ketoglutarate

oxalosuccinate

pyruvate acetyl-CoA

alanine

aspartate

glutamine

phenylalanine

phenylalanine

acetoacetyl-CoA

citrateoxaloacetate

fumarate

succinate

succinyl-CoA

malate isocitrate

aconitate

α-ketoglutarate

oxalosuccinate

pyruvate acetyl-CoA acetoacetyl-CoA

ketonesglucosegluconeogenesis ketogenesis

The α-keto acids derived from catabolism of many amino acids are intermediates in glycolysis and the citric acid cycle.

The α-keto acids derived from catabolism of some amino acids are broken down to acetyl-CoA or acetoacetyl-CoA and are

oxidized by the citric acid cycle or converted to ketone bodies.

Amino acids that can be converted into pyruvate, α-ketoglutarate, succinyl-CoA, fumarate, and oxaloacetate can be converted into glucose by gluconeogenesis and are said to be glucogenic.

These α-keto acids may, therefore, replenish citric acid cycle intermediates.

These amino acids are said to be ketogenic.

Glucogenic and Ketogenic Amino Acids

citrate

fumarate

succinate

succinyl-CoA

malate isocitrate

aconitate

α-ketoglutarate

oxalosuccinate

pyruvate acetyl-CoA acetoacetyl-CoA ketones

gluconeogenesis

alanine glycine threonine cysteine serine

aspartate asparagine

glutamate glutamine arginine histidine

proline isoleucine methionine

valine threonine

aspartate phenylalanine

tyrosine

leucine lysine phenylalanine tyrosine

tryptophan

leucine isoleucine tryptophan threonine

Glucogenic and Ketogenic Amino Acids

oxaloacetate

Overview of Amino Acid Catabolism

H3N C C

H

RO

OThe catabolism of amino acids takes

place in three stages:

1) Removal of the amino group, leaving the carbon

skeleton of the amino acid.

2) Breakdown of the carbon skeletons to a glycolytic

intermediate, citric acid cycle intermediate, or acetyl-S-CoA.

3) Oxidation of these intermediates to CO2 and H2O

with the production of ATP.

small pieces

CO2 H2O

ATP

The Fate of the Amino Group of Amino Acids

The extraction of the amino group from amino acids must be done in such a way so as not to increase

blood ammonium ion levels above normal values.

NH4+

The normal concentration of NH4+ ion in the blood is 3.0 x 10-5 to 6.0 x 10-5 M.

Nitrogen is present in the bloodstream in the form of ammonium ions:

Above these concentrations (hyperammonemia) coma may result.

NH4+ NH3

R-NH2R-NH3+

Because they are weak bases, amines in high concentrations cannot be used to transport nitrogen

atoms in the bloodstream.

C

CO O

H3N H

CH2CH2C

O O

C

CO O

H3N H

CH2CH2C

O NH2L-glutamate

C

CO O

H3N H

CH2CH2C

O NH2

L-glutamine

C

CO O

CH3

H3N H

L-alanine

“Neutral” forms of nitrogen atoms are used for

transport in the bloodstream and across

membranes.

The pathway for the extraction of amino groups from amino acids consists of three phases:

1) Conversion of amino groups from all amino acids into a single product, glutamate, by transamination.

2) Conversion of glutamate into α-ketoglutarate by oxidative deamination, releasing NH4+ .

3) Conversion of NH4+ into urea, which is extracted from the blood by the kidneys and excreted.

The Fate of the Amino Group of Amino Acids

CHC

O

O

CHC

O

O

CHC

O

O

CH2C

O

O NH3

aspartate

fumarate

α-amino acid α-keto acid

α-keto glutarate glutamate

NAD+, (NADP)+,

H2O

NADH, (NADPH),

NH4+ CO2

Urea cycleH2N

CNH2

O

Urea

transamination

oxidative deamination

Transamination

alpha amino acid alpha keto acid

HC

CO O

R1

NH3 C

CO O

R2

O C

CO O

R1

O HC

CO O

R2

NH3++HC

CO O

R1

NH3 C

CO O

R2

O C

CO O

R1

O HC

CO O

R2

NH3++

The first step in amino acid metabolism is the removal of the amino group by transamination followed by oxidative deamination.

A transamination reaction can be represented as:

Transamination

Amino acid1 + α-ketoacid2 ⇔ α-ketoacid1 + amino acid 2

HC

CO O

R1

NH3 C

CO O

R2

O C

CO O

R1

O HC

CO O

R2

NH3++

Transamination reactions occur in all cells.

The enzymes responsible for transaminations are called transaminases or amino transferases.

Most transaminases are specific for α-ketoglutarate but are less specific for the amino acid.

This means that the amino groups of almost all amino acids end up on glutamic acid (i.e., L-glutamate).

L-amino acid + α-ketoglutarate ⇔ α-ketoacid + L-glutamate

C

CO O

R

H3N C

CO O

CH2

O C

CO O

R

O C

CO O

CH2

H3N++H

CH2

CO O

CH2

CO O

H

Transamination

One exception to this rule is in skeletal muscle, where transaminases use pyruvate as the amino acceptor, producing alanine as the product.

L-amino acid + pyruvate ⇔ α-ketoacid + L-alanine

C

CO O

R

H3N C

CO O

CH3

O C

CO O

R

O C

CO O

CH3

H3N++H H

Another example of a specific transaminase reaction is aspartate transaminase:

L-aspartate + α-ketoglutarate ⇔ oxaloacetate + L-glutamate

C

CO O

CH2

H3N C

CO O

CH2

O C

CO O

O C

CO O

H3N++H H

CO O

CH2

CO O

CH2

CH2

CO O

CH2

CO O

Transamination in Diagnostic Laboratory Medicine

The presence of alanine transaminase (glutamate:pyruvate transaminase or GPT) and aspartate transaminase

(glutamate:oxaloacetate transaminase, or GOT) in the bloodstream, above a certain base level, may indicate liver damage.

Serum GPT and GOT (SGPT and SGOT) tests measure the

severity and stage of liver damage.

Vitamin B-6 as a Coenzyme

All transaminases require the coenzyme pyridoxal phosphate (derived from pyridoxine, vitamin B-6):

N

C

HO

H3C

CH2

O H

H

O P

O

O

O

pyridoxal phosphate

Vitamin preparations may contain the precursor to pyridoxal phosphate in different forms:

N

C

HO

H3C

CH2OH

O H

HN

CH2NH2

HO

H3C

CH2OH

H

N

CH2OH

HO

H3C

CH2OH

H

pyridoxalpyridoxamine

pyridoxine

The Transaminase Mechanism

N

C

HO

H3C

CH2

N H

H

O PO32-

CHCO

O

R

CC

O O

RH3N H

H2O

CC

O O

RO

H2O

pyridoxamine phosphate

N

C

HO

H3C

CH2

N H

H

O PO32-

CCOO

R

H

N

CHO

H3C

CH2

O H

H

O PO32-

N

CHO

H3C

CH2

H3N

H

O PO32-

H

H

pyridoxal phosphate

alpha amino acid

alpha keto acid

The Fate of the Amino Group of Amino Acids

NH4+

Nitrogen is present in the bloodstream in the form of ammonium ions:

The pathway for the extraction of amino groups from amino acids consists of three phases:

1) Conversion of amino groups from all amino acids into a single product, glutamate, by transamination.

2) Conversion of glutamate into α-ketoglutarate by oxidative deamination, releasing NH4+ .

3) Conversion of NH4+ into urea, which is extracted from the blood by the kidneys and excreted.

The Fate of the Amino Group of Amino Acids

CHC

O

O

CHC

O

O

CHC

O

O

CH2C

O

O NH3

aspartate

fumarate

α-amino acid α-keto acid

α-keto glutarate glutamate

CO2

Urea cycleH2N

CNH2

O

Urea

transamination

oxidative deamination

NAD+, (NADP)+,

H2O

NADH, (NADPH),

NH4+

C

CO O

CH2

O

CH2C

O O

C

CO O

H3N H

CH2CH2C

O O

NH4++

NADH (NADPH)NAD+ (NADP+) H2O

glutamate dehydrogenase

Oxidative Deamination

This reaction is reversible and provides a mechanism for

1) generating ammonium ion for excretion as urea

2) generating a-ketoglutarate

3) assimilating ammonium ion for use in other metabolic pathways in the liver and kidneys

glutamate α-ketoglutarate

DOES NOT ENTER THE

BLOODSTREAM

Amino Group and Ammonia Transport

Amino groups collected in extrahepatic tissues in the form of glutamate must be packaged in a non-toxic form for

transport through the blood to the liver.

Glutamate, itself, cannot pass through the cell membranes.

Two different transport forms are used:

1) Production of glutamine in most cell types

2) Production of alanine in muscle cells.

Glutamine Production

+ NH4+ + ATP + H+ + ADP +Pi

C

CO O

H3N H

CH2CH2C

O O

C

CO O

H3N H

CH2CH2C

O NH2

glutamine synthetase

In almost all cell types, glutamine synthetase catalyzes the formation of glutamine from glutamate:

The glutamine, thus formed, is electrically neutral, nontoxic, and can pass through the cell membranes into the blood. The concentration of glutamine in the blood is higher than any other amino acid.

L-glutamate L-glutamine

Amino Group and Ammonia Transport

Once in the liver, the reverse reaction takes place and glutamine is deaminated to ammonium ion and glutamate.

C

CO O

H3N H

CH2CH2C

O NH2

C

CO O

H3N H

CH2CH2C

O O

+ H2O + NH4+

Glutamate can then be oxidatively deaminated to ammonium ion and α-ketoglutarate.

L-glutamate L-glutamine

C

CO O

H3N H

CH2CH2C

O O

L-glutamate

C

CO O

CH2

O

CH2C

O O

α-ketoglutarate

+ NAD+ + H2O + NADH + NH4+

glutamate dehydrogenase

L-glutamate

C

CO O

H3N H

CH2CH2C

O O

C

CO O

CH2

O

CH2C

O O

α-ketoglutarate

+ NADP+ + H2O+ NADPH + NH4+

glutamate dehydrogenase

In active muscle cells, large quantities of ammonium ion are produced. After two reactions, alanine is formed. Like glutamine, alanine is

electrically neutral:

C

CO O

CH2

OC

CO O

H3N H

CH2C

O O

CH2CH2C

O O

C

CO O

CH3

O C

CO O

CH3

H3N++ H

α-ketoglutarateL-glutamate

pyruvate alanine

C

CO O

CH3

H3N H

C

CO O

H3N H

CH2CH2C

O NH2

Like glutamine, alanine is electrically neutral and readily traverses membranes and enters the blood stream.

alanine

L-glutamine

In the liver, the combination of transamination and oxidative deamination reaction releases ammonium ions.

Glucose

Pyruvate Lactate

2 ATPGlucose

Pyruvate

Alanine

Urea

6 ATP

4 ATP

Alanine

𝛂-amino acid

𝛂-keto acid

Liver Muscle tissue

N

Glucose/Alanine Cycle

Overall Reaction:

NH4+ + HCO3- + 3 ATP + aspartate (-NH3+)

urea + 2 ADP + AMP + 4 PO43- + fumarate

Urea Cycle

H2NC

NH2

O

CNH2H2N

O

C

C

O O

HH3N

CH2CH2CH2NH

COH2N

C

C

O O

HH3N

CH2CH2CH2NH3

C

C

O O

HH3N

CH2CH2CH2NH

CNH2H2N

C

C

O O

HH3N

CH2CH2CH2NH

CNH2N C

CO O

H

CH2C

O O

arginine

ornithine

citrulline

arginosuccinate

urea

Urea Cycle

ornithine

arginine

argininosuccinate

citrulline

citrulline

ornithine

urea

glutamate

glutamate a-ketoglutarate

NH4+

Pi

mitochondria

cytosolaspartate

fumarate

ATP

AMP, 2Pi

Urea Cycle

carbamoyl phosphate

HCO3-2 ATP

2 ADP,Pi

Urea Cycle

CNH2H2N

O

C

C

O O

HH3N

CH2CH2CH2NH

COH2N

C

C

O O

HH3N

CH2CH2CH2NH3

C

C

O O

HH3N

CH2CH2CH2NH

CNH2H2N

C

C

O O

HH3N

CH2CH2CH2NH

CNH2N C

CO O

H

CH2C

O O

arginine

ornithine

citrulline

arginosuccinate

urea

C

CO O

H

CH2

CO O

H3N

C

C

H

H

C

O

O

C O

O

aspartate

fumarate

COH2N

P O

O

O

O

P O

O

O

NH4+ , HCO3-, 2ATP

2ADP,Pi

carbamoyl phosphate

Urea Cycle

AMP,2Pi ATP

Overall Reaction:

NH4+ + HCO3- + 3 ATP + aspartate

urea + 2 ADP + AMP + 4 PO43- + fumarate

Urea Cycle

ATP, via TCA

cycle

Glucose

Fatty acids

Ketones

Urea

NH4+

Amino acid pool

Dietary protein pyruvate,

acetyl-CoA, acetoacetate,

TCA cycle intermediates

Liver proteins Plasma proteins

Other nitrogen-containing

compounds

aquatic invertebrates,

bony fishes, crocodiles

mammals, sharks, some bony fishes,

turtles

birds, insects, reptiles, land

gastropodsscorpions,

spiders

NH3H2N

C O

H2N

CHN

CNH

C

C

NH

C

HN

O

O

O

Nitrogen excretion products for various organisms

Water solubility

Energy needed to produce

N→?

CHN

CN

C

C

NH

CH

N

H2N

O

ammonia urea uric acid guanine

Habitat

H2N

C O

H2N

NH3

H2N

C O

H2N

NH3